Mutations in the PEX (also known as PHEX) gene are responsible for X-linked hypophosphatemic rickets (HYP). To gain insight into the role of PEX in normal physiology we have cloned the human full-length cDNA and studied its tissue expression, subcellular localization, and peptidase activity. We show that the cDNA encodes a 749 amino acid protein structurally related to a family of neutral endopeptidases that include neprilysin (NEP) as prototype. By Northern blot analysis, the size of the full-length PEX transcript is 6.5 kb. PEX expression, as determined by semi-quantitative PCR, is high in bone and in tumor tissue associated with the paraneoplastic syndrome of renal phosphate wasting. PEX is glycosylated in the presence of canine microsomal membranes and partitions exclusively in the detergent phase from Triton X-114 extractions of transiently transfected COS cells. Immunofluorescence studies in A293 cells expressing PEX tagged with a c-myc epitope show a predominant cell-surface location for the protein with its C-terminal domain in the extracellular compartment, substantiating the assumption that PEX, like other members of the neutral endopeptidase family, is a type II integral membrane glycoprotein. Cell membranes from cultured COS cells transiently expressing PEX efficiently degrade exogenously added PTH-derived peptides, demonstrating for the first time that recombinant PEX can function as an endopeptidase. PEX peptidase activity may provide a convenient target for pharmacological intervention in states of altered phosphate homeostasis and in metabolic bone diseases.
X-linked hypophosphatemic rickets (HYP) is the most common inherited disorder of renal phosphate wasting characterized by severe hypophosphatemia, renal phosphate wasting, reduced serum concentrations of 1,25-dihydroxyvitamin D levels, and defective bone mineralization. Until recently, much of our understanding of HYP has been facilitated by the availability of two murine homologues, the Hyp and Gy mice, which exhibit many of the phenotypic features of HYP. Through positional cloning, however, a gene which spans the deleted region Xp22.1 in HYP patients, or is mutated in non-deletion patients with the disorder, was identified (designated PEX) and its partial cDNA sequence reported (The HYP Consortium (1995) Nature Genetics 11, 130-136). The predicted human PEX gene product, as well as its murine homologue (Du, L. et al. (1996) Genomics 36, 22-28), exhibit homology to a family of neutral endopeptidases involved in either activation or degradation of a number of peptide hormones. It has been postulated that PEX metabolizes a peptide hormone that modulates renal tubular phosphate handling. Such an activity could involve either the processing of a phosphate-reabsorbing hormone precursor to its active form or the inactivation of a circulating phosphaturic factor. These speculations notwithstanding, the physiologic function of the PEX gene product and the mechanisms that lead to the renal and skeletal abnormalities of HYP remain to be defined.
Oncogenous hypophosphatemic osteomalacia (OHO) is a rare acquired disorder of phosphate homeostasis with biochemical and physical abnormalities similar to HYP. This syndrome is associated with a variety of histologically distinct, usually benign, mesenchymal tumors whose excision promptly reverses the metabolic abnormalities and results in cure of the bone disease. It is generally thought that a factor(s) produced by these tumors promotes phosphaturia and inhibits the renal conversion of 25-hydroxyvitamin D to 1,25-dihydroxyvitamin D. The nature of the phosphaturic substance remains unknown and is likely distinct from both parathyroid hormone (PTH) and calcitonin, two polypeptide hormones known to inhibit the renal tubular reabsorption of phosphorus. Because of the striking similarity in the clinical presentation of patients with OHO and HYP, it is postulated that the factor causing phosphaturia in OHO is the active form of the PEX substrate. The identification and characterization of the putative PEX substrate, referred to as phosphatonin, however, will require first a better understanding of PEX function.
To date, there is still a need to understand how local factors produced in the bone regulate bone formation and bone resorption. Derangement of these factors leads to metabolic bone diseases. Pharmacological manipulation of such factors may serve as a novel approach to the treatment of these disorders.
It would be highly desirable to be provided with a tool in the treatment of metabolic bone diseases, such as osteomalacia and osteoporosis.